Tripartite_analysis_manuscript

1 Growth Analysis for Moss and Cyanobacteria

1.1 Introduction

The symbiosis between Sphagnum mosses and cyanobacteria has long been recognized (Limpricht, 1890) and has been implicated in improved moss growth. This together with evidence form other plant - cyanobacteria symbioses suggest that this is not just a symbiosis (interacting species), but perhaps a mutualsitc interaction – thereby benefiting both interacting speices. To investigate this further, Colleen Hulsey and Megan Gable conducted a numer of experiments to investigate the growth consequences of cyanobacteria and sphagnum symbiosis on growth patterns at differing pH. conditions.

1.1.1 Growth 2018 symbiosis approach and measurements

*Experimental conditions & design +conducted on 08/09/2018 +plants pH nitrogen depleted for 2 weeks prior to start +experiment ran for 2 weeks +condition 1: Nitrogen (N), 6 replicates -N, 2 replicates for low N (5mL/L NaNO3– Mm?) +Condition 2: pH, 3.5, 5.5, 8.5 +speices: Sphagnum fallax MN alone, Nostoc muscorum 1037 alone, both species together +amounts: 1 moss capitulum, 0.1g cyanobacteria +3 - 12 well plates, 2 ml liquid medium per well.

*Measurements +fluorcam: area_mm2, QY_max (initial, weekly, final) +pH initial, pH weekly, pH final +weight beginning and end after water removed with centrifugation

*tests +simple scatter and boxplots to view realtionship

1.1.2 Growth 2018 data import and summary

library(tidyverse)
#knitr by default will set the working directory to the source file location
#setwd("~/Documents/Manuscripts:data/Symbiosis_study_2018")
dat1 <- read_csv("fallax_cyano_symbiosis_colleen_10_01_2018.csv") %>% mutate(delta_weight = delta_weight_g *1000)
names(dat1)

## [1] "Sample_well"        "component_measured" "condition"         
## [4] "initial_pH"         "final_pH"           "initial_weight_g"  
## [7] "final_weight_g"     "delta_weight_g"     "delta_weight"

summary(dat1)

##  Sample_well        component_measured  condition           initial_pH   
##  Length:72          Length:72          Length:72          Min.   :3.500  
##  Class :character   Class :character   Class :character   1st Qu.:3.500  
##  Mode  :character   Mode  :character   Mode  :character   Median :5.500  
##                                                           Mean   :5.833  
##                                                           3rd Qu.:8.500  
##                                                           Max.   :8.500  
##                                                                          
##     final_pH     initial_weight_g  final_weight_g    delta_weight_g    
##  Min.   :3.700   Min.   :0.00140   Min.   :0.01430   Min.   :-0.03560  
##  1st Qu.:5.800   1st Qu.:0.01342   1st Qu.:0.04083   1st Qu.: 0.01082  
##  Median :6.200   Median :0.09345   Median :0.09570   Median : 0.03030  
##  Mean   :6.117   Mean   :0.06677   Mean   :0.10127   Mean   : 0.03766  
##  3rd Qu.:6.700   3rd Qu.:0.10622   3rd Qu.:0.15368   3rd Qu.: 0.06940  
##  Max.   :7.440   Max.   :0.12040   Max.   :0.22060   Max.   : 0.11090  
##  NA's   :12      NA's   :12        NA's   :12                          
##   delta_weight   
##  Min.   :-35.60  
##  1st Qu.: 10.82  
##  Median : 30.30  
##  Mean   : 37.66  
##  3rd Qu.: 69.40  
##  Max.   :110.90  
## 

dim(dat1)

## [1] 72  9

1.1.3 Growth 2018 data boxplot analysis

1. Delta growth (weight) by organism regardless of pH; 2. Delta growth by organism and by pH; 3. Delta growth for entier wel by pH

Fig. 1: 2018 data, change in growth (initial weight (wt) - final wt among the two symbiotic partners alone and when added togather but grown seperately (_seperate) and when grown together in same well (_symbiosis). Cyano in symbiosis includes, endophytes, epiphytes and free living cyano. Data incuded all pH conditions

Fig. 2: 2018 data, change in growth among interacting partners faceted by starting pH condiiton. Measured components as described in fig. 1

Fig. 3: 2018 data, total well biomass change from experiment start to finish. Includes moss + cyano endophyte and free living cyanobacteria not attached to plant

## Warning: Removed 12 rows containing non-finite values (stat_boxplot).

## Warning: Removed 12 rows containing missing values (geom_point).

Fig. 4: 2018 data, change in pH change as measured by (final - initial values)

## Warning: Removed 12 rows containing non-finite values (stat_boxplot).

## Warning: Removed 12 rows containing missing values (geom_point).

Fig. 5: 2018 data, absolute value of pH change as measured by (final - initial values)

1.2 Growth 2017 symbiosis approach and measurements

need experimental details here…

1.2.1 Growth 2017 data import and summary

dat2 <- read_csv("fallax_cyano_symbiosis_allyssa_colleen_2017.csv")
dat3 <- dat2 %>% filter(pH == 3.5 | pH == 5.5 | pH == 8.5)
names(dat3)

## [1] "condition"      "replicate"      "pH"             "plate"         
## [5] "well"           "weight_initial" "weight_final"   "delta_weight"

summary(dat3)

##   condition           replicate         pH            plate   
##  Length:72          Min.   :1.0   Min.   :3.500   Min.   :1   
##  Class :character   1st Qu.:2.0   1st Qu.:3.500   1st Qu.:3   
##  Mode  :character   Median :3.5   Median :5.500   Median :5   
##                     Mean   :3.5   Mean   :5.833   Mean   :5   
##                     3rd Qu.:5.0   3rd Qu.:8.500   3rd Qu.:7   
##                     Max.   :6.0   Max.   :8.500   Max.   :9   
##                     NA's   :18                    NA's   :18  
##       well        weight_initial   weight_final     delta_weight    
##  Min.   : 1.000   Min.   : 3.11   Min.   :  8.67   Min.   :-21.330  
##  1st Qu.: 3.000   1st Qu.:21.73   1st Qu.: 29.20   1st Qu.:  8.562  
##  Median : 6.500   Median :30.00   Median : 46.91   Median : 23.870  
##  Mean   : 6.333   Mean   :33.72   Mean   : 64.98   Mean   : 32.358  
##  3rd Qu.: 9.000   3rd Qu.:40.02   3rd Qu.: 96.84   3rd Qu.: 46.220  
##  Max.   :12.000   Max.   :87.22   Max.   :177.43   Max.   :127.830  
##  NA's   :18       NA's   :18      NA's   :18

dim(dat3)

## [1] 72  8

1.2.2 Boxplot analysis of 2017 data

Fig. 6: 2017 data, change in growth (initial weight (wt) - final wt among the two symbiotic partners alone, when added togather but grown seperately (_seperate) and when grown together in same well (_symbiosis). Conditions aggregated over pH

Fig. 7: 2017 data, change in growth (initial weight (wt) - final wt among the two symbiotic partners alone, when added togather but grown seperately (_seperate) and when grown together in same well (_symbiosis). Similar to Fig. 6 except faceted by pH

Fig. 8: Combined 2018 and 2017 data for change in growth (initial weight (wt) - among the two symbiotic partners alone, when added togather but grown seperately (_seperate) and when grown together in same well (_symbiosis). Data is faceted by pH

1.2.3 ANOVA analysis of 2017 and 2018 data.

Specifically testing if cyano and moss grown alone, increased in growth at a different rate than when grown together in smbiosis.

##              Df Sum Sq Mean Sq F value   Pr(>F)    
## condition     3  20854    6951   9.251 4.05e-05 ***
## pH            2   8810    4405   5.862  0.00473 ** 
## condition:pH  6   2464     411   0.546  0.77075    
## Residuals    60  45086     751                     
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

computing summary stats

## # A tibble: 12 x 5
## # Groups:   condition [?]
##    condition                         pH     count  mean    sd
##    <fct>                             <fct>  <int> <dbl> <dbl>
##  1 cyano_alone_2017                  pH_3.5     6  1.38  9.17
##  2 cyano_alone_2017                  pH_5.5     6  5.64 16.4 
##  3 cyano_alone_2017                  pH_8.5     6 29.8  39.4 
##  4 moss_alone_2017                   pH_3.5     6 13.8  14.0 
##  5 moss_alone_2017                   pH_5.5     6 32.1  26.2 
##  6 moss_alone_2017                   pH_8.5     6 24.2  15.4 
##  7 Cyanobacteria_moss_seperate_2017  pH_3.5     6 15.2   5.52
##  8 Cyanobacteria_moss_seperate_2017  pH_5.5     6 37.7  37.7 
##  9 Cyanobacteria_moss_seperate_2017  pH_8.5     6 54.0  51.4 
## 10 Cyanobacteria_moss_symbiosis_2017 pH_3.5     6 44.1  20.1 
## 11 Cyanobacteria_moss_symbiosis_2017 pH_5.5     6 55.5  23.3 
## 12 Cyanobacteria_moss_symbiosis_2017 pH_8.5     6 74.8  30.7

mean seperation iwth Tukeys

##   Tukey multiple comparisons of means
##     95% family-wise confidence level
## 
## Fit: aov(formula = delta_weight ~ condition + pH + condition:pH, data = dat.2017)
## 
## $condition
##                                                                        diff
## moss_alone_2017-cyano_alone_2017                                   11.11278
## Cyanobacteria_moss_seperate_2017-cyano_alone_2017                  23.38444
## Cyanobacteria_moss_symbiosis_2017-cyano_alone_2017                 45.84889
## Cyanobacteria_moss_seperate_2017-moss_alone_2017                   12.27167
## Cyanobacteria_moss_symbiosis_2017-moss_alone_2017                  34.73611
## Cyanobacteria_moss_symbiosis_2017-Cyanobacteria_moss_seperate_2017 22.46444
##                                                                           lwr
## moss_alone_2017-cyano_alone_2017                                   -13.033058
## Cyanobacteria_moss_seperate_2017-cyano_alone_2017                   -0.761391
## Cyanobacteria_moss_symbiosis_2017-cyano_alone_2017                  21.703053
## Cyanobacteria_moss_seperate_2017-moss_alone_2017                   -11.874169
## Cyanobacteria_moss_symbiosis_2017-moss_alone_2017                   10.590276
## Cyanobacteria_moss_symbiosis_2017-Cyanobacteria_moss_seperate_2017  -1.681391
##                                                                         upr
## moss_alone_2017-cyano_alone_2017                                   35.25861
## Cyanobacteria_moss_seperate_2017-cyano_alone_2017                  47.53028
## Cyanobacteria_moss_symbiosis_2017-cyano_alone_2017                 69.99472
## Cyanobacteria_moss_seperate_2017-moss_alone_2017                   36.41750
## Cyanobacteria_moss_symbiosis_2017-moss_alone_2017                  58.88195
## Cyanobacteria_moss_symbiosis_2017-Cyanobacteria_moss_seperate_2017 46.61028
##                                                                        p adj
## moss_alone_2017-cyano_alone_2017                                   0.6191255
## Cyanobacteria_moss_seperate_2017-cyano_alone_2017                  0.0610500
## Cyanobacteria_moss_symbiosis_2017-cyano_alone_2017                 0.0000289
## Cyanobacteria_moss_seperate_2017-moss_alone_2017                   0.5396394
## Cyanobacteria_moss_symbiosis_2017-moss_alone_2017                  0.0018777
## Cyanobacteria_moss_symbiosis_2017-Cyanobacteria_moss_seperate_2017 0.0771172

resdiual check

## 
##  Shapiro-Wilk normality test
## 
## data:  aov_residuals
## W = 0.96343, p-value = 0.03459

Starting the 2018 data

##              Df Sum Sq Mean Sq F value   Pr(>F)    
## condition     3  11694    3898   5.216  0.00429 ** 
## pH            2  32248   16124  21.577 6.93e-07 ***
## condition:pH  6  11846    1974   2.642  0.03159 *  
## Residuals    36  26902     747                     
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

computing summary stats for 2018

## # A tibble: 12 x 5
## # Groups:   condition [?]
##    condition                         pH     count   mean    sd
##    <fct>                             <fct>  <int>  <dbl> <dbl>
##  1 moss_alone_2018                   pH_3.5     4   32.6 31.4 
##  2 moss_alone_2018                   pH_5.5     4   35.5 16.4 
##  3 moss_alone_2018                   pH_8.5     4   29.8 11.5 
##  4 cyano_alone_2018                  pH_3.5     4  -21.4 24.9 
##  5 cyano_alone_2018                  pH_5.5     4   20.5 28.6 
##  6 cyano_alone_2018                  pH_8.5     4   63.2 23.0 
##  7 Cyanobacteria_moss_symbiosis_2018 pH_3.5     4   14.3 21.5 
##  8 Cyanobacteria_moss_symbiosis_2018 pH_5.5     4   59.6 42.4 
##  9 Cyanobacteria_moss_symbiosis_2018 pH_8.5     4  105.   5.81
## 10 Cyanobacteria_moss_seperate_2018  pH_3.5     4   11.3 41.3 
## 11 Cyanobacteria_moss_seperate_2018  pH_5.5     4   56.0 36.3 
## 12 Cyanobacteria_moss_seperate_2018  pH_8.5     4   93.0 16.9

mean seperation with Tukeys for 2018

##   Tukey multiple comparisons of means
##     95% family-wise confidence level
## 
## Fit: aov(formula = delta_weight ~ condition + pH + condition:pH, data = dat.2018)
## 
## $condition
##                                                                          diff
## cyano_alone_2018-moss_alone_2018                                   -11.858333
## Cyanobacteria_moss_symbiosis_2018-moss_alone_2018                   26.866667
## Cyanobacteria_moss_seperate_2018-moss_alone_2018                    20.800000
## Cyanobacteria_moss_symbiosis_2018-cyano_alone_2018                  38.725000
## Cyanobacteria_moss_seperate_2018-cyano_alone_2018                   32.658333
## Cyanobacteria_moss_seperate_2018-Cyanobacteria_moss_symbiosis_2018  -6.066667
##                                                                           lwr
## cyano_alone_2018-moss_alone_2018                                   -41.915110
## Cyanobacteria_moss_symbiosis_2018-moss_alone_2018                   -3.190110
## Cyanobacteria_moss_seperate_2018-moss_alone_2018                    -9.256777
## Cyanobacteria_moss_symbiosis_2018-cyano_alone_2018                   8.668223
## Cyanobacteria_moss_seperate_2018-cyano_alone_2018                    2.601556
## Cyanobacteria_moss_seperate_2018-Cyanobacteria_moss_symbiosis_2018 -36.123444
##                                                                         upr
## cyano_alone_2018-moss_alone_2018                                   18.19844
## Cyanobacteria_moss_symbiosis_2018-moss_alone_2018                  56.92344
## Cyanobacteria_moss_seperate_2018-moss_alone_2018                   50.85678
## Cyanobacteria_moss_symbiosis_2018-cyano_alone_2018                 68.78178
## Cyanobacteria_moss_seperate_2018-cyano_alone_2018                  62.71511
## Cyanobacteria_moss_seperate_2018-Cyanobacteria_moss_symbiosis_2018 23.99011
##                                                                        p adj
## cyano_alone_2018-moss_alone_2018                                   0.7140984
## Cyanobacteria_moss_symbiosis_2018-moss_alone_2018                  0.0937375
## Cyanobacteria_moss_seperate_2018-moss_alone_2018                   0.2614148
## Cyanobacteria_moss_symbiosis_2018-cyano_alone_2018                 0.0071606
## Cyanobacteria_moss_seperate_2018-cyano_alone_2018                  0.0288313
## Cyanobacteria_moss_seperate_2018-Cyanobacteria_moss_symbiosis_2018 0.9476697

resdiual check 2018 data

Shapiro-Wilk test for normality

## 
##  Shapiro-Wilk normality test
## 
## data:  aov_residuals
## W = 0.96343, p-value = 0.03459

#Levene's Test for homogenity of variances

library(car)

## Loading required package: carData

## 
## Attaching package: 'car'

## The following object is masked from 'package:dplyr':
## 
##     recode

## The following object is masked from 'package:purrr':
## 
##     some

leveneTest(delta_weight ~ condition, data = dat.2018)

## Levene's Test for Homogeneity of Variance (center = median)
##       Df F value  Pr(>F)  
## group  3  3.7312 0.01787 *
##       44                  
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

leveneTest(delta_weight ~ pH, data = dat.2018)

## Levene's Test for Homogeneity of Variance (center = median)
##       Df F value Pr(>F)
## group  2  0.0209 0.9793
##       45

2 Metabolite Analysis

2.1 Introduction to design

(in progress)

2.2 metablite change when cyanobacteria pellet is crossfed with moss supernantant

Goal – determine significant changes in metabolites between cyanobacteria and moss using kruskal wallis test.

To look at how suernatant from 1) cyano pellet when incubated with moss supernatant (Ph2-super-Nmusc-SfallaxSupp) compares to 2) moss only supernantant (Ph1-super-Sfallax).

Header- crossfed cyano pellet with moss supernatant

p.value	met_direction	compound_atlas_ID	label	Identified_metabolite
0.0138744	uptake_up by cyano	0130_NADpositive_11p55	NAD	NAD
0.0138744	eluted_by cyano	0001_4-Pyridoxic_acid_positive_1p32	4-PYRIDOXATE	4-PYRIDOXATE
0.0138744	uptake_up by cyano	0207_UDP-glucose_negative_11p47	URIDINE 5’-DIPHOSPHOGLUCOSE	URIDINE 5’-DIPHOSPHOGLUCOSE / URIDINE 5’-DIPHOSPHOGALACTOSE
0.0138744	uptake_up by cyano	0208_UDP-D-glucose_negative_11p52	URIDINE 5’-DIPHOSPHOGALACTOSE	URIDINE 5’-DIPHOSPHOGLUCOSE / URIDINE 5’-DIPHOSPHOGALACTOSE
0.0179605	eluted_by cyano	0056_2-Hydroxy-4methylthio_butyric_acid_negative_3p66	2-HYDROXY-4-(METHYLTHIO)BUTYRIC ACID	2-HYDROXY-4-(METHYLTHIO)BUTYRIC ACID
0.0179605	eluted_by cyano	0223_L-histidine_negative_13p06	L-histidine	L-histidine
0.0201646	eluted_by cyano	0216_L-histidine_negative_11p85	L-HISTIDINE	L-HISTIDINE
0.0201646	uptake_up by cyano	0230_L-ornithine_negative_18p98	L-ORNITHINE	L-ORNITHINE

## [1] "# elluted cyanobacteria metabolites (p.value <= 0.05) is: 65"

## [1] "# taken cyanobacteria metabolites (p.value <= 0.05) is: 79"

2.3 metablite change when moss pellet is crossfed with cyanobacteria supernantant

Goal – determine significant changes in metabolites between cyanobacteria and moss using kruskal wallis test.

Supernatant changes from 1) suernatant from moss pellet when incubated with cyanobacteria supernatant (Ph2-super-Sfallax-NmuscSupp) compared to 2) cyanobacteria pellet only supernantant (Ph1-super-Nmusc).

Header- crossfed moss pellet with cyanobacteria supernatant

p.value	met_direction	compound_atlas_ID	label	Identified_metabolite
0.0138744	uptake_up by moss	0105_cytidine_negative_6p43	CYTIDINE	CYTIDINE
0.0138744	uptake_up by moss	0174_thiamine_negative_10p28	THIAMINE	THIAMINE
0.0138744	eluted_by moss	0226_ANSERINE_negative_14p47	L-ANSERINE	L-ANSERINE
0.0138744	uptake_up by moss	0056_2-Hydroxy-4methylthio_butyric_acid_negative_3p66	2-HYDROXY-4-(METHYLTHIO)BUTYRIC ACID	2-HYDROXY-4-(METHYLTHIO)BUTYRIC ACID
0.0179605	uptake_up by moss	0133_xanthosine_negative_8p42	XANTHOSINE	XANTHOSINE
0.0179605	eluted_by moss	0058_DFructose_positive_7p00	fructose	fructose
0.0201646	eluted_by moss	0130_NADpositive_11p55	NAD	NAD
0.0201646	uptake_up by moss	0150_2-Phospho-D-Glyceric_Acid_positive_12p51	L-2-PHOSPHOGLYCERIC ACID	L-2-PHOSPHOGLYCERIC ACID

## [1] "# elluted moss metabolites (p.value <= 0.05) is: 103"

## [1] "# taken moss metabolites (p.value <= 0.05) is: 44"

2.4 metablite change when fungus pellet is crossfed with cyanobacteria supernantant

Goal – determine significant changes in metabolites between fungus and cyanobacteria using kruskal wallis test.

Supernatant changes from 1) suernatant from fungus pellet when incubated with cyanobacteria supernatant (Ph2-super-Tacro-NmuscSupp) compared to 2) cyanobacteria pellet only supernantant (Ph1-super-Nmusc).

Header- crossfed fungus pellet with cyano supernatant

p.value	met_direction	compound_atlas_ID	label	Identified_metabolite
0.0138744	uptake_up by fungus	0001_4-Pyridoxic_acid_positive_1p32	4-PYRIDOXATE	4-PYRIDOXATE
0.0138744	uptake_up by fungus	0011_1-Methyladenosine_positive_2p17	1-METHYLADENOSINE	1-METHYLADENOSINE
0.0138744	eluted_by fungus	0218_D-Glycerol_1-phosphate_negative_12p21	SN-GLYCEROL 3-PHOSPHATE	SN-GLYCEROL 3-PHOSPHATE
0.0138744	eluted_by fungus	0142_beta-Glycerophosphoric_acid_positive_11p81	GLYCEROL 2-PHOSPHATE	GLYCEROL 2-PHOSPHATE / SN-GLYCEROL 3-PHOSPHATE
0.0179605	uptake_up by fungus	0082_N-Acetyl-DL-methionine_negative_5p11	N-ACETYL-DL-METHIONINE	N-ACETYL-DL-METHIONINE
0.0179605	uptake_up by fungus	0074_3-METHOXY-L-TYROSINE_positive_8p17	3-METHOXY-L-TYROSINE	3-METHOXY-L-TYROSINE
0.0179605	uptake_up by fungus	0005_1-Hydroxy-2-naphthoic_acid_negative_1p14	1-HYDROXY-2-NAPHTHOATE	1-HYDROXY-2-NAPHTHOATE
0.0201646	uptake_up by fungus	0150_2-Phospho-D-Glyceric_Acid_positive_12p51	L-2-PHOSPHOGLYCERIC ACID	L-2-PHOSPHOGLYCERIC ACID

## [1] "# elluted fungal metabolites (p.value <= 0.05) is: 51"

## [1] "# taken fungal metabolites (p.value <= 0.05) is: 87"

2.5 metablite change when fungus pellet is crossfed with moss supernantant

Goal – determine significant changes in metabolites between fungus and moss using kruskal wallis test.

Supernatant changes from 1) suernatant from fungus pellet when incubated with moss supernatant (Ph2-super-Tacro-SfallaxSupp) compared to 2) moss pellet only supernantant (Ph1-super-Sfallax).

Header- crossfed fungus pellet with moss supernatant

p.value	met_direction	compound_atlas_ID	label	Identified_metabolite
0.0138744	uptake_up by fungus	0005_dihydrouracil_positive_1p57	5,6-DIHYDROURACIL	5,6-DIHYDROURACIL
0.0138744	eluted_by fungus	0001_4-Pyridoxic_acid_positive_1p32	4-PYRIDOXATE	4-PYRIDOXATE
0.0138744	eluted_by fungus	0174_thiamine_negative_10p28	THIAMINE	THIAMINE
0.0138744	uptake_up by fungus	0095_2deoxyguanosine_negative_6p18	2’-DEOXYGUANOSINE	2’-DEOXYGUANOSINE
0.0138744	uptake_up by fungus	0083_DIPALMITOYL_CEPHALIN_negative_5p15	1,2-DIPALMITOYL-RAC-GLYCERO-3-PHOSPHOETHANOLAMINE	1,2-DIPALMITOYL-RAC-GLYCERO-3-PHOSPHOETHANOLAMINE
0.0179605	eluted_by fungus	0102_Orotic_acid_negative_6p35	OROTATE	OROTATE
0.0179605	eluted_by fungus	0056_2-Hydroxy-4methylthio_butyric_acid_negative_3p66	2-HYDROXY-4-(METHYLTHIO)BUTYRIC ACID	2-HYDROXY-4-(METHYLTHIO)BUTYRIC ACID
0.0201646	uptake_up by fungus	0011_1-Methyladenosine_positive_2p17	1-METHYLADENOSINE	1-METHYLADENOSINE

## [1] "# elluted fungal metabolites (p.value <= 0.05) is: 118"

## [1] "# taken fungal metabolites (p.value <= 0.05) is: 32"

2.6 metablite change when moss pellet is crossfed with fungus supernantant

Goal – determine significant changes in metabolites between fungus and moss using kruskal wallis test.

Supernatant changes from 1) suernatant from moss pellet when incubated with fungus supernatant (Ph2-super-Sfallax-TacroSupp) compared to 2) fungus pellet only supernantant (Ph1-super-Tacro).

Header- crossfed moss pellet with fungus supernatant

p.value	met_direction	compound_atlas_ID	label	Identified_metabolite
0.0138744	eluted_by moss	0132_3-Nitro-L-tyrosine_negative_8p33	3-NITRO-L-TYROSINE	3-NITRO-L-TYROSINE
0.0138744	eluted_by moss	0146_AICA_ribonucleotide_positive_12p02	5-AMINOIMIDAZOLE-4-CARBOXAMIDE-1-BETA-D-RIBOFURANOSYL 5’-MONOPHOSPHATE	5-AMINOIMIDAZOLE-4-CARBOXAMIDE-1-BETA-D-RIBOFURANOSYL 5’-MONOPHOSPHATE
0.0138744	eluted_by moss	0105_S-adenosyl-L-homocysteine_positive_10p36	S-(5’-ADENOSYL)-L-HOMOCYSTEINE	S-(5’-ADENOSYL)-L-HOMOCYSTEINE
0.0138744	eluted_by moss	0134_3-Hydroxy-dl-kynurenine_negative_8p42	3-HYDROXYKYNURENINE	3-HYDROXYKYNURENINE
0.0179605	eluted_by moss	0043_2DEOXYURIDINE_negative_2p47	DEOXYURIDINE	DEOXYURIDINE
0.0179605	eluted_by moss	0069_5-Hydroxyindole-3-acetic_acid_positive_7p96	5-HYDROXYINDOLEACETATE	5-HYDROXYINDOLEACETATE
0.0201646	eluted_by moss	0130_NADpositive_11p55	NAD	NAD
0.0201646	uptake_up by moss	0162_N-Acetylneuraminic_Acid_negative_9p99	N-ACETYLNEURAMINATE	N-ACETYLNEURAMINATE

## [1] "# elluted fungal metabolites (p.value <= 0.05) is: 102"

## [1] "# taken fungal metabolites (p.value <= 0.05) is: 21"

2.7 metablite change when cyano pellet is crossfed with fungus supernantant

Supernatant changes from 1) suernatant from cyano pellet when incubated with fungus supernatant (Ph2-super-Nmusc-TacroSupp) compared to 2) fungus pellet only supernantant (Ph1-super-Tacro).

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Header- crossfed moss pellet with fungus supernatant

p.value	met_direction	compound_atlas_ID	label	Identified_metabolite
0.0138744	eluted_by cyano	0133_xanthosine_negative_8p42	XANTHOSINE	XANTHOSINE
0.0138744	eluted_by cyano	0132_3-Nitro-L-tyrosine_negative_8p33	3-NITRO-L-TYROSINE	3-NITRO-L-TYROSINE
0.0138744	eluted_by cyano	0105_S-adenosyl-L-homocysteine_positive_10p36	S-(5’-ADENOSYL)-L-HOMOCYSTEINE	S-(5’-ADENOSYL)-L-HOMOCYSTEINE
0.0138744	eluted_by cyano	0009_5deoxyadenosine_positive_2p03	5’-DEOXYADENOSINE	5’-DEOXYADENOSINE / 2’-DEOXYADENOSINE
0.0138744	eluted_by cyano	0013_2deoxyadenosine_positive_2p42	2’-DEOXYADENOSINE	5’-DEOXYADENOSINE / 2’-DEOXYADENOSINE
0.0138744	eluted_by cyano	0074_3-METHOXY-L-TYROSINE_positive_8p17	3-METHOXY-L-TYROSINE	3-METHOXY-L-TYROSINE
0.0138744	eluted_by cyano	0223_L-histidine_negative_13p06	L-histidine	L-histidine
0.0179605	eluted_by cyano	0069_5-Hydroxyindole-3-acetic_acid_positive_7p96	5-HYDROXYINDOLEACETATE	5-HYDROXYINDOLEACETATE

## [1] "# elluted cyano metabolites (p.value <= 0.05) is: 125"

## [1] "# taken cyano metabolites (p.value <= 0.05) is: 25"